Broad challenge area: (06) Enabling technologies Specific challenge topic: 06-OD-106 Renewable affinity reagents for epigenomic research. The goals of this proposal are (i) to establish a technology platform for generating high-quality, well- validated synthetic antibodies and (ii) to produce the initial set of synthetic antibodies to high-value antigens useful for epigenetic research. Synthetic antibodies are generated using state-of-the-art in vitro techniques without animal immunization. They are monoclonal by definition and their expression clones, together with their immunochemical properties, are permanently archived and easily distributed. Together, these attributes make synthetic antibodies much superior alternatives to monoclonal and polyclonal antibodies. Affinity reagents to epigenetic markers such as histone tails containing specific post-translational modifications (PTMs) and to other proteins involved in epigenetic regulation are critically important tools in epigenomics research. They are the central component of the chromatin immunoprecipitation (ChIP), the standard, powerful tool and also essential for immunohistochemical and biochemical analyses of protein complexes involved in regulating and responding to epigenomic changes. It is now painfully clear that conventional antibodies present severe limitations. Monoclonal antibody production is slow and expensive, and there is little control on their affinity or selectivity. Polyclonal antibodies have a fundamental problem in reproducibility and specificity, which amount to unacceptable levels of experimental variations. This situation represents a critical gap in our ability to perform epigenomic profiling in a highly quality-controlled manner. Our team includes three leaders in the field of synthetic antibody engineering and a leader in proteomic analysis of histone modifications. We have collectively developed a complete spectrum of fully recombinant antibody technologies whose performance exceeds that of natural antibodies. In this project, we will combine our complementary expertise to further enhance and apply these powerful technologies for generating synthetic antibodies of exquisite specificity and affinity to histone tail PTMs and other proteins involved in epigenetic regulation. The specific aims of this project are (1) to develop an integrated technological platform for the production and characterization of synthetic antibodies with exquisite antigen binding properties;(2) to generate synthetic antibodies to histone tail peptides and to polycomb components;(3) to rigorously validate generated antibodies for ChIP applications using multitude of analytical tools;and (4) to develop a Web- database and logistical infrastructure for antibody distribution. Renewable, synthetic antibodies generated in this project will make an immediate and strong impact on epigenomics research, because they will be highly validated and their properties (affinity and specificity) will not change. This project will employ and train additional personnel in the rapidly growing field of recombinant antibodies. Together, this project perfectly aligns with the general goal of the ARRA challenge grants and with the specific challenge of generating renewable affinity reagents for epigenomic research. Subtle modifications (post-translational modifications) in the chemical composition of histones are critically important biomarkers in the epigenomic regulation of transcription, a process that is critical for the understanding of cell development and the onset of disease states. Research in this field has been severely hindered by a lack of high- quality "affinity reagents" that can selectively detect and capture histone modifications and proteins associated with them. This project will establish a state-of-the-art platform and generate high-quality, renewable affinity reagents for epigenomic targets, thereby eliminating a major bottleneck in epigenomic research.